G-protein coupled receptors (GPCRs) are proteins responsible for transducing a signal within a cell. GPCRs have usually seven transmembrane domains. Upon binding of a ligand to an extra-cellular portion or fragment of a GPCR, a signal is transduced within the cell that results in a change in a biological or physiological property or behaviour of the cell. GPCRs, along with G-proteins and effectors (intracellular enzymes and channels modulated by G-proteins), are the components of a modular signalling system that connects the state of intra-cellular second messengers to extra-cellular inputs.
GPCR genes and gene products can modulate various physiological processes and are potential causative agents of disease. The GPCRs seem to be of critical importance to both the central nervous system and peripheral physiological processes.
The GPCR protein superfamily is represented in five families: Family I, receptors typified by rhodopsin and the beta2-adrenergic receptor and currently represented by over 200 unique members; Family II, the parathyroid hormone/calcitonin/secretin receptor family; Family III, the metabotropic glutamate receptor family, Family IV, the CAMP receptor family, important in the chemotaxis and development of D. discoideum; and Family V, the fungal mating pheromone receptor such as STE2.
G proteins represent a family of heterotrimeric proteins composed of α, β and γ subunits, that bind guanine nucleotides. These proteins are usually linked to cell surface receptors (receptors containing seven transmembrane domains) for signal transduction. Indeed, following ligand binding to the GPCR, a conformational change is transmitted to the G protein, which causes the α-subunit to exchange a bound GDP molecule for a GTP molecule and to dissociate from the βγ-subunits.
The GTP-bound form of the α, β and γ-subunits typically functions as an effector-modulating moiety, leading to the production of second messengers, such as cAMP (e.g. by activation of adenyl cyclase), diacylglycerol or inositol phosphates.
Greater than 20 different types of α-subunits are known in humans. These subunits associate with a small pool of β and γ subunits. Examples of mammalian G proteins include Gi, Go, Gq, Gs and Gt. G proteins are described extensively in Lodish et al., Molecular Cell Biology (Scientific American Books Inc., New York, N.Y., 1995; and also by Downes and Gautam, 1999, The G-Protein Subunit Gene Families. Genomics 62:544-552), the contents of both of which are incorporated herein by reference.
Known and uncharacterized GPCRs currently constitute major targets for drug action and development. There are ongoing efforts to identify new G protein coupled receptors which can be used to screen for new agonists and antagonists having potential prophylactic and therapeutic properties.
More than 300 GPCRs have been cloned to date, excluding the family of olfactory receptors. Mechanistically, approximately 50-60% of all clinically relevant drugs act by modulating the functions of various GPCRs (Cudermann et al., J. Mol. Med., 73:51-63, 1995).
GPR43 is a member of the rhodopsin like receptors family, cloned in 1997. It shows a homology of 38% with another orphan GPCR, GPR41 and 27% with transmembrane domains of mouse PAR1 receptor. The gene encoding GPR43 coding gene is located on human chromosome 19q31 (Sawzdargo et al., 1997). GPR43 has been described as a gene induced by IL-9 in mouse cytokine dependent T helper cell lines and bone marrow derived primary mast cells. In addition, GPR43 mRNA transcription is stimulated in the lung, intestine and stomach of transgenic mice overexpressing IL-9. GPR43 mRNA is also induced in splenoytes by mitogens, such as concanavalin A, and this induction is blocked by aminosterol compounds (see WO99/15656). GPR43 polynucleotide and amino acid sequences are disclosed in U.S. Pat. Nos. 5,910,430 and 6,180,365B1 and in WO00/28083, WO98/40483, WO99/15656 and WO00/22129, each of which is incorporated herein by reference.
Short chain fatty acids (SCFA) include but are not limited to acetate, propionate, butyrate and valerate. SCFA are produced by microbial fermentation in the hindgut in considerable amounts. Most of the anions in hindgut contents are SCFA, mainly acetate, propionate and butyrate. SCFA are rapidly absorbed, and the total SCFA concentration in peripheral blood reaches 79 μM (Cummings, 1987). Among the different SCFAs, acetate is the principal anion and can also be produced in different tissues by biochemical synthesis (Bergman, 1990). Acetate is present in the plasma at a concentration of 59 to 85 μM and its concentration can be increased by 20 fold after ethanol administration (Lundquist et al., 1960). It is believed that most plasma acetate is derived from the splanchnic bed and is used by other tissues where it can account for almost 7% of basal energy expenditure. Butyrate is produced by bacterial fermentation of dietary fibers in the colon lumen, and dramatically affects the proliferation of colon cancer cells in vitro experiments. Various periodontal and root canal pathogens, such as the Bacteroides species, can produce significant amounts of short chain fatty acids. (SCFA). Short-chain fatty acids are also physiological regulators of growth and differentiation in the gastrointestinal tract and can act as antibacterial agents. There is some evidence that SCFA metabolism is involved in the development of colitis ulcerosa, diverticulosis and colorectal cancer. The differences between the effects of SCFA on cell proliferation, differentiation and apoptosis of colonocytes in vivo and in vitro indicate that in addition to direct effects of SCFA, systemic effects such as neural and humoral factors are also of crucial importance. The opposing effects of SCFA on proliferation and apoptosis in normal colonocytes and in colon cancer cells demonstrate possibilities for prevention and/or therapy of colonic diseases.